Heparin Induces Rat Aorta Relaxation via Integrin-Dependent Activation of Muscarinic M3 Receptors

• Published in: Hypertension (Dallas, Tex. 1979) Tex. 1979), 2010-10, Vol.56 (4), p.713-721
• Main Authors: Paredes-Gamero, Edgar J, Medeiros, Valquíria P, Farias, Eduardo H.C, Justo, Giselle Z, Trindade, Edvaldo S, Andrade-Lopes, Ana L, Godinho, Rosely O, de Miranda, Antonio, Ferreira, Alice T, Tersariol, Ivarne L.S, Nader, Helena B
• Format: Article
• Language: English
• Subjects: Acetylcholine - pharmacology; Animals; Anticoagulants - pharmacology; Aorta, Thoracic - drug effects; Aorta, Thoracic - metabolism; Aorta, Thoracic - physiology; Arterial hypertension. Arterial hypotension; Atropine - pharmacology; Biological and medical sciences; Blood and lymphatic vessels; Blotting, Western; Cardiology. Vascular system; Cattle; Cell Line; Dose-Response Relationship, Drug; Endothelium, Vascular - metabolism; Endothelium, Vascular - physiology; Experimental diseases; Fluorescence Resonance Energy Transfer; Heparin - pharmacology; In Vitro Techniques; Integrins - metabolism; Male; Medical sciences; Nitric Oxide - biosynthesis; Oligopeptides - pharmacology; Paxillin - metabolism; Protein Binding; Proto-Oncogene Proteins pp60(c-src) - metabolism; Rats; Rats, Wistar; Receptor, Muscarinic M3 - metabolism; Vasodilation - drug effects; Vasodilator Agents - pharmacology
• ISSN: 0194-911X; 1524-4563
• DOI: 10.1161/HYPERTENSIONAHA.110.156877

Previous reports have shown that heparin may promote human hypotension and vascular relaxation by elevation of NO levels through unclear mechanisms. We hypothesized that endothelial muscarinic M3 receptor activation mediates the heparin-induced vasodilation of rat aortic rings. The experiments were carried out using unfractionated heparin extracted from bovine intestinal mucosa, which elicited an endothelium and NO-dependent relaxation of aortic segments with maximal potency and efficacy (EC50100±10 μmol/L; Emax41±3%). Atropine and 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide inhibitors reduced the heparin-dependent relaxation, indicating that M3 muscarinic receptor is involved in this phenomenon. However, no direct binding of heparin to muscarinic receptors was observed. More importantly, studies performed using the arginine-glycine-aspartic acid peptide and 1-(1,1-dimethylethyl)-3-(1-naphthalenyl)-1H-pyrazolo[3,4-day]pyrimidin-4-amine, an Src family inhibitor, reduced by 51% and 73% the heparin-dependent relaxation, respectively, suggesting the coupling of heparin and M3 receptor through extracellular matrix molecules and integrin. Furthermore, unfractionated heparin induced activation of focal adhesion protein kinase, Src, and paxillin. Finally, fluorescence resonance energy transfer approach confirmed the interaction of the M3 receptor to integrin. Taken together, these data demonstrate the participation of M3 receptor and integrin in heparin-dependent relaxation of vascular smooth muscle. These results provide new insights into the molecular mechanism and potential pharmacological action of heparin in vascular physiology.